title: Crucial role for DNA ligase III in mitochondria but not in Xrcc1-dependent repair.
abstract: Mammalian cells have three ATP-dependent DNA ligases, which are required for DNA replication and repair. Homologues of ligase I (Lig1) and ligase IV (Lig4) are ubiquitous in Eukarya, whereas ligase III (Lig3), which has nuclear and mitochondrial forms, appears to be restricted to vertebrates. Lig3 is implicated in various DNA repair pathways with its partner protein Xrcc1 (ref. 1). Deletion of Lig3 results in early embryonic lethality in mice, as well as apparent cellular lethality, which has precluded definitive characterization of Lig3 function. Here we used pre-emptive complementation to determine the viability requirement for Lig3 in mammalian cells and its requirement in DNA repair. Various forms of Lig3 were introduced stably into mouse embryonic stem (mES) cells containing a conditional allele of Lig3 that could be deleted with Cre recombinase. With this approach, we find that the mitochondrial, but not nuclear, Lig3 is required for cellular viability. Although the catalytic function of Lig3 is required, the zinc finger (ZnF) and BRCA1 carboxy (C)-terminal-related (BRCT) domains of Lig3 are not. Remarkably, the viability requirement for Lig3 can be circumvented by targeting Lig1 to the mitochondria or expressing Chlorella virus DNA ligase, the minimal eukaryal nick-sealing enzyme, or Escherichia coli LigA, an NAD(+)-dependent ligase. Lig3-null cells are not sensitive to several DNA-damaging agents that sensitize Xrcc1-deficient cells. Our results establish a role for Lig3 in mitochondria, but distinguish it from its interacting protein Xrcc1.
authors.affiliation: Developmental Biology Program, Memorial Sloan-Kettering Cancer Center, New York, New York 10065, USA.
authors.forename: Deniz
authors.initials: D
authors.lastname: Simsek
authors.forename: Amy
authors.initials: A
authors.lastname: Furda
authors.forename: Yankun
authors.initials: Y
authors.lastname: Gao
authors.forename: Jérôme
authors.initials: J
authors.lastname: Artus
authors.forename: Erika
authors.initials: E
authors.lastname: Brunet
authors.forename: Anna-Katerina
authors.initials: AK
authors.lastname: Hadjantonakis
authors.forename: Bennett
authors.initials: B
authors.lastname: Van Houten
authors.forename: Stewart
authors.initials: S
authors.lastname: Shuman
authors.forename: Peter J
authors.initials: PJ
authors.lastname: McKinnon
authors.forename: Maria
authors.initials: M
authors.lastname: Jasin
mesh_terms: D000818:Animals; D055162:Biocatalysis; D002470:Cell Survival; D004249:DNA Damage; D000072481:DNA Ligase ATP; D011088:DNA Ligases / Q000737:chemistry / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D004260:DNA Repair; D004272:DNA, Mitochondrial / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D053595:Embryonic Stem Cells / Q000378:metabolism; D020043:Genes, Essential; D005816:Genetic Complementation Test; D006801:Humans; D051379:Mice; D008928:Mitochondria / Q000201:enzymology* / Q000235:genetics* / Q000473:pathology; D000075223:Poly-ADP-Ribose Binding Proteins; D017434:Protein Structure, Tertiary; D012854:Sister Chromatid Exchange / Q000187:drug effects; D000076105:X-ray Repair Cross Complementing Protein 1; D029867:Xenopus Proteins
chemical_list: D004272:DNA, Mitochondrial; D004268:DNA-Binding Proteins; D000075223:Poly-ADP-Ribose Binding Proteins; D000076105:X-ray Repair Cross Complementing Protein 1; C000619371:XRCC1 protein, human; D029867:Xenopus Proteins; C000619352:Xrcc1 protein, mouse; D011088:DNA Ligases; D000072481:DNA Ligase ATP; C000618816:DNA ligase III alpha protein, Xenopus; C000618817:LIG3 protein, human; C000618818:Lig3 protein, mouse